We propose advanced development and validation of novel technology that will maximize the quality and utility of biologic fluid specimens, and permit the measurement of previously invisible low abundance biomarkers. The technology is transformative and paradigm-shifting because it immediately and economically solves fundamental roadblocks paralyzing the body fluid cancer biomarker field. In a single step, in minutes, our technology overcomes the severe problems of biomarker preservation, low abundance, and masking by unwanted proteins. The technology is novel porous, buoyant, core-shell hydrogel nanoparticles containing high affinity reactive chemical baits that harvests biomarkers in body fluids. Our nanoparticles can be pre loaded into Vacutainer(R) blood collection tubes, or other body fluid collection vessels. Upon contact with the sample, the suspended nanoparticles immediately affinity-sequester target biomarkers inside the particles, exclude albumin, fully protect the biomarkers from degradation (even at elevated temperatures), and massively concentrate the sequestered biomarkers into a small volume. The technology can dramatically (demonstrated up to 10,000 fold) improve the lower limits of detection and the precision of: a) mass spectrometry (MS) biomarker discovery, b) quantitation by multiple reaction monitoring (MRM), or c) quantification by any clinical grade immunoassay. All of the Aims and Milestones of the predicate IMAT NCI R21 CA137706 grant have been exceeded. We have discovered more than a dozen novel chemical baits with preferential high affinities (KD <10-11 M) for specific low abundance protein analytes. We discovered a novel shell chemistry that selectively prevented unwanted entry of all size albumin-derived peptides without hindering the penetration of non-albumin small proteins and peptides. Labile body fluid biomarkers, that would otherwise rapidly degrade, are completely preserved at the time of collection, thus obviating the need for costly freezing or proteinase inhibitors. Low abundant proteins previously invisible to discovery by MS, and previously not measureable by MRM, or clinical immunoassays, can now be quantified with high precision within the linear range of the assay. Our technology is completely innovative, as documented by 3 allowed (2 issued) patents that have been commercially licensed, and 14 publications. No existing technology can solve all of the aforementioned roadblocks, and attain a capture/elution efficiency near 100%. The technology has recently permitted the MS identification of new serum and plasma proteins not listed in the international HUPO database. Under the proposed Aims we will scale up the technology to conduct blind validation of its performance precision, accuracy, improved detection sensitivity, and preservation capacity, in two large (n = 400 sera and n = 74 plasma) well-controlled clinical sera/plasma sample sets (64 analytes). We will extend the technology to urine and sweat to open up these biofluids as a new category for biomarker research, using innovative approaches to solve fundamental problems of volume, perishability, and low protein concentration for these biofluids.